Applied Immune Sciences has developed a polystyrene device based cell separation technology for therapeutic application in infectious diseases, cancer and autoimmune disease. It is the long term goal of AIS to combine this technology with efficient gene transfer technology to develop novel therapeutics for commercialization. To achieve this goal we have chosen to develop recombinant adeno-associated virus (rAAV) vectors for high efficiency gene transduction into targeted cell types. Because of the nonpathogenic properties of AAV, and the ability of this virus to infect both dividing and nondividing cells, AAV is a good candidate for a multipurpose, safe, DNA transduction system. The specific aims of the proposed research is to: l) develop a system to very simply produce large amounts of high titer stocks; 2) optimize the infection efficiency of rAAV stocks in a variety of human cells of therapeutic interest; 3) develop methods to purify the rAAV; 4) investigate the stability and integrity of rAAV; and 5) pilot scale up production of rAAV. The proposed work will take advantage of mammalian cell episomal vectors, recombinant transducing viruses and AIS' cell separation technology to develop simple systems for safe genetic engineering of specific cell types.